As the miniaturization of analytical instruments continuously proceeds [M. V. Novotny and D. Ishii, Eds., "Microcolumn Separations", Elsevier, Amsterdam, 1985; R. P. W. Scott, Ed., "Small-Bore Liquid Chromatography Columns: Their Properties and Uses", John Wiley & Sons, New York, Chem. Anal., 72, 1984; and Y. Hirata, J. Microcol. Sep., 2 (1990) 214], the packed capillary, also known as microbore, columns with an internal diameter (I.D.) of from 10 to 1000 .mu.m are used increasingly in microseparation techniques, such as microcolumn high performance liquid chromatography (micro-HPLC), supercritical fluid chromatography (SFC) [K. M. Payne, I. L. Davies, K. D. Bartle, K. E. Markides and M. L. Lee, J. Chromatogr., 477 (1989) 161], gas chromatography (GC) [F. J. Yang, HRC & CC, 6 (1983) 348], electrochromatography (EC) [C. Yan, D. Schaufelberger and F. Erni,] and multidimensional chromatography [K. D. Bartle, I. L. Davies, M. W. Raynor, A. A. Clifford and J. P. Kithinji, J. Microcol. Sep., 1 (1989) 63]. The increasing demands of packed capillary columns justify the need for searching for more efficient and more economical column-packing technology. Although many efforts have been made in this direction, the slurry packing technique [R. P. W. Scott, Ed., "Small-Bore Liquid Chromatography Columns: Their Properties and Uses", John Wiley & Sons, New York, Chem. Anal., Vol. 72, 1984; and Y. Hirata, J. Microcol. Sep., 2 (1990) 214], so far, has been employed almost exclusively for packing capillary columns. A few reports on a dry packing method [G. Crescentini, F. Bruner, F. Mangani and G. Yafeng, Anal. Chem., 60, (1988) 1659; and J. H. Knox and I. H. Grant, Chromatographia, 32 (1991) 317] have also been made.
In the slurry packing method, a suspension of packing material stored in a reservoir is pushed into a column under high pressure (generally from about 200 to 500 atm.). This method is useful for packing capillary columns with fairly good efficiency. However, the involved technique has a number of shortcomings and limitations. One particular disadvantage is that slurry packing is limited to packing particles having a size larger than one micron. Packing of a capillary column with particles of a smaller particle size is important for further improvement in the efficiency of microseparation techniques, especially in electrochromatography. It has been predicted theoretically, and proved experimentally, that the electroosmotic flow is essentially unaffected by a particle size down to at least 1.5 .mu.m. As the particle size is reduced below 1 .mu.m, the plate heights are dominated mainly by the B-term (axial diffusion) in the van Deemter equation. ##EQU1## Based on the preceding, an efficiency of a million theoretical plates in a time of less than 30 minutes could be achievable in electrochromatography.
However, to pack such a column efficiently with fine particles is a real challenge since, conventionally with slurry packing, extremely high pressure is required. Especially when wide diameter capillaries with a thin wall are being packed, application of such high pressure could be dangerous. Therefore, alternative ways must be found to pack micron and submicron particles in order to improve further the performance of EC in particular, and microseparation in general. In addition, the slurry packing of capillary columns is rather tedious and time consuming because each capillary must be packed individually. Furthermore, the particles (having any particular size distribution) are pushed into a column in a non-selective manner so that the homogeneity of a column packed by the slurry method is rather low.
Electrokinetic separation is known and has developed rapidly since 1981 [Y. Lee, "Capillary Electrophoreses: Principles, practice and applications", Elsevier, Amsterdam, J Chromatogr. Library, Vol. 52, 1992; J. Vindevogel and P. Sandra, "Introduction to Micellar Electrokinetic Chromatography", Huthig Heidelberg, 1992; J. W. Jorgenson, K. D. Lukacs, Anal Chem., 53 (1981) 1298]. In addition to separation and analysis of compounds, capillary zone electrophoresis (CZE) has been used for characterization and separation of particulate materials, such as submicron polystyrene spheres [B. B. van Orman, G. L. Mcintyre, Am. Lab., Nov., Vol. 66, 1990] and viral particles [S. Hjeren, K. Elenbring, F. Kilar, J. L. Liao, A. J. C. Chen; C. J. Siebert and M. D. Zhu, J. Chromatogr., 403 (1987) 47], as well as colloidal silica sols with diameters in the range of from 5 to 500 nm [R. M. McCormick, J. Liquid Chromatogr., 14 (1991) 939].
During the past three decades, silica-based material has been the dominating stationary phase for high performance liquid chromatography (HPLC). However, it has a serious disadvantage when an alkaline condition is presented because of its limited pH stability above pH 8. What makes things worse is that when positively charged species, e.g., organic bases or certain proteins, are dealt with under the limited alkaline conditions, the interaction of solutes with a negatively charged silica surface (pH &gt;2) causes severe tailing. The effect of the restricted pH stability of a silica matrix is even more pronounced in electrochromatography where a higher pH value usually is desirable for higher mobile phase velocity, which depends on the surface charge of the stationary phase particles.
An aluminum oxide matrix provides an alternative to silica due to its inherent higher pH stability, generally within a pH range of from 2 to 13 [J. J. Pesek, J. E. Sandoval and M. Su, J. Chromatogr., 630 (1993) 95; K. Cabrera, D. Lubda and G. Jung, Kontakte (Darmstadt), 1 (1992) 32]. Depending on the pH value of the mobile phase, the chromatographic properties can be changed due to the amphoteric character of aluminum oxide [K. Cabrera, D. Lubda and G. Jung, Kontakte (Darmstadt), 1 (1992) 32]. Particularly in electrochromatography, a column packed with an aluminum oxide-based stationary phase is useful as a multi-purpose column since the sign of the surface charge on the particles, and consequently the direction of electroosmotic flow (EOF) can be altered by varying the pH value of the mobile phase.